Methods of using (1s,3s)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid

ABSTRACT

(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid also known as CPP-115 can be used to treat addiction and neurological disorders without side effects such as visual field defects caused by vigabatrin (Sabril).

PRIORITY CLAIM

This application is a Continuation patent application of U.S.application Ser. No. 13/581,187, filed on Aug. 24, 2012, now allowed,which is a U.S. National Phase application under 35 U.S.C. §371 ofInternational Patent Application No. PCT/US11/26309, filed on Feb. 25,2011, which claims the benefit of U.S. Provisional Application No.61/308,030 filed on Feb. 25, 2010, the contents of which are expresslyincorporated by reference. All references cited herein are expresslyincorporated by reference.

GOVERNMENT SUPPORT

This invention was made with government support under GM066132, awardedby the National Institutes of Health, and DE-ACO2-98CH10886 andDE-SC0012704 awarded by the Department of Energy. The government hascertain rights in the invention.

Vigabatrin (γ-vinyl GABA) is sold worldwide under the trademark Sabrilfor treatment of epilepsy and has been studied for treatment of drugaddiction. Vigabatrin's well-known mechanism of action is theirreversible inhibition of gamma-aminobutyric acid-aminotransferase(GABA-AT). This enzyme is responsible for the catabolism of gammaaminobutyric acid (GABA) in the brain. Inhibition of this enzyme resultsin an elevation of brain levels of GABA. The elevation of brain GABA(the brain's primary inhibitory neurotransmitter) results in a decreaseof neuron excitability and as such reduces uncontrolled firing ofneurons, which leads to a reduction in epileptic seizures.

Unfortunately, long term use of the drug results in a constriction ofthe patient's visual field which in turn has prevented vigabatrin fromgaining widespread usage. Visual field defects were detectable in somepatients in less than 2 months after initiation of therapy and was mostpronounced at about 1 year. One third or more of patients were affectedwith visual field defects after multiple years of therapy withvigabatrin. In the United States, the Food and Drug Administrationdeemed Vigabatrin unapprovable in 1998 as a direct result of the visualfield defects following agency conclusions that “FDA unaware of way toreliably prevent damage” and “FDA unable to propose sound monitoringplan” to identify damage. Vigabatrin was subsequently approved fortreatment of spasms in infants and epileptic seizures in 2009. The FDA'spress release on the approval stated:

“Damage to vision is an important safety concern with the use of Sabril.The drug will have a boxed warning to alert health care professionals tothis risk of a progressive loss of peripheral vision with potentialdecrease in visual acuity. The risk of vision damage may increase basedon the dosage and duration of use, but even the lowest doses of Sabrilcan cause vision damage. Periodic vision testing is required for thosetaking Sabril. Because of the risk of permanent vision damage, the drugwill be available only through a restricted distribution program.”

As launched Sabril contains a black boxed warning as follows:

WARNING: VISION LOSS See full prescribing information for complete boxedwarning SABRIL causes progressive and permanent bilateral concentricvisual field constriction in a high percentage of patients. In somecases, SABRIL may also reduce visual acuity. Risk increases with totaldose and duration of use, but no exposure to SABRIL is known that isfree of risk of vision loss Risk of new and worsening vision losscontinues as long as SABRIL is used, and possibly after discontinuingSABRIL Periodic vision testing is required for patients on SABRIL, butcannot reliably prevent vision damage Because of the risk of permanentvision loss, SABRIL is available only through a special restricteddistribution program

U.S. Pat. No. 6,713,497 teaches that vitamin B6 may be used to mitigatevisual field defects caused by vigabatrin. Taurine deficiency is alsoknown in the art as a possible contributing factor to the visual fielddefects resulting from vigabatrin administration. Jammoul, et al.,Taurine Deficiency is a Cause of Vigabatrin Induced RetinalPhototoxicity, Ann. Neurol 2009: 65:98-107.

A need exists in the art to treat patients with GABA aminotransferaseinhibitors without the side effects of vigabatrin.

U.S. Pat. Nos. 7,381,748 and 6,794,413, which are incorporated herein byreference disclose the compound(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid. Theliterature has shown that(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid isapproximately 186 times more potent as a mechanism-based inactivator ofγ-aminobutyric acid aminotransferase (GABA-AT) than the anticonvulsantdrug and GABA-AT inactivator vigabatrin (1, Sabril™) under nonoptimalconditions (Pan, Y.; Qiu, J.; Silverman, R. B. Design, Synthesis, andBiological Activity of a Difluoro-substituted, Conformationally-rigidVigabatrin Analogue As a Potent γ-Aminobutyric Acid AminotransferaseInhibitor. J. Med. Chem. 2003, 46, 5292-5293).

It has been surprisingly discovered that(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid does notinhibit [³H]GABA uptake in neurons, astrocytes, or mammalian cellsrecombinantly expressing the four different human GABA transportersubtypes (hGAT-1, hBGT-1, hGAT-2, and hGAT-3), nor does it bind toGABA_(A) or GABA_(B) receptors in rat brain homogenate, or affectGABA_(C) receptor activity in Xenopus laevis oocytes. Thus, it appearsthat (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid isselective for GABA-AT.

U.S. Pat. Nos. 6,906,099; 6,890,951; 6,828,349; 6,593,367; 6,541,520;6,395,783; 6,323,239; and 6,057,368, describe and/or claim the use ofvigabatrin in the treatment of addiction from cocaine, nicotine,methamphetamine, morphine, heroin, ethanol, phencyclidine,methylenedioxymethamphetamine, and/or PCT. The contents of such patentsare expressly incorporated herein by reference.

U.S. Pat. No. 6,462,084 describes and/or claims the use of vigabatrin inthe treatment of obsessive compulsive disorders including generalanxiety disorder, pathological or compulsive gambling disorder,compulsive eating (obesity), body dysmorphic disorder, hypochondriasis,pathologic grooming conditions, kleptomania, pyromania, attentiondeficit hyperactivity disorder and impulse control disorders. Thecontents of U.S. Pat. No. 6,462,084 is expressly incorporated herein byreference.

U.S. Pat. No. 6,939,876 describes and/or claims the use of vigabatrin inthe treatment to prevent addiction to opioid analgesics by coadministration of vigabatrin. The contents of U.S. Pat. No. 6,939,876 isexpressly incorporated herein by reference.

Gabaergic drugs are those that improve secretion or transmission ofGABA. These drugs as a family have been used to treat a wide variety ofnervous system disorders including fibromyalgia, neuropathy, migrainesrelated to epilepsy, restless leg syndrome, and post traumatic distressdisorder. Gabaergic drugs include GABA_(A) and GABA_(B) receptorligands, GABA reuptake inhibitors, GABA aminotransferase inhibitors,GABA analogs, or molecules containing GABA itself. Preferred GABAergicdrugs include valproate and its derivatives, vigabatrin, pregabalin,gabapentin and tiagabine.

As reported in the literature, although vigabatrin is an irreversibleinhibitor of GABA-AT, its binding to GABA-AT is relatively weak (K₁=3.2mM, k_(inact)=0.37, k_(inact)/K₁=0.11)¹ Pan, Yue; Qiu, Jian; Silverman,Richard B.; “Design, Synthesis, and biological Activity of aDifluoro-Substituted, Conformationally rigid Vigabatrin Analogue as aPotent γ-Aminobutyric Acid Aminotransferase Inhibitor”, J. Med. Chem.,2003, 46(25), 5292-5293. Dr. Richard Silverman elucidated the mechanismby which vigabatrin inactivates GABA-AT. Burke, James R.; Silverman,Richard B.; “Mechanism of inactivation of γ-aminobutyric acidaminotransferase by the antiepilepsy drug γ-vinyl GABA (vigabatrin)”, J.Am. Chem. Soc., 1991, 113(24), 9341-9349 and then set out to develop anew GABA-AT inhibitor that would exhibit superior binding and enzymeinactivation when compared to vigabatrin. The development workultimately culminated in the development of(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid (U.S. Pat.Nos. 6,794,413 and 7,381,748, referred to as compound 2 in the textbelow). The contents of U.S. Pat. Nos. 6,794,413 and 7,381,748 areexpressly incorporated herein by reference. During this developmentprocess, several other candidate compounds were created, including(1R,4S)-4-amino-cyclopent-2-ene-1-carboxylic acid (compound(1R,4S)-(+)-3 in reference 3 and referred to as compound 1 in the textbelow) and (1S,3S)-3-amino-4-methylenyl-1-cyclopentanoic acid (compound6 in reference 1). As published in 2003, Silverman, et al. (Pan, Yue;Qiu, Jian; Silverman, Richard B.; “Design, Synthesis, and biologicalActivity of a Difluoro-Substituted, Conformationally rigid VigabatrinAnalogue as a Potent γ-Aminobutyric Acid Aminotransferase Inhibitor”, J.Med. Chem., 2003, 46(25), 5292-5293) stated that compound 1 “was not aGABA-AT inactivator but was a very good substrate with a specificityconstant almost five times greater than that of GABA.” It was furtherimplied that compound 1's failure to inhibit GABA-AT made it a poorcandidate for further development as an antiepileptic medication andwork proceeded on a new candidate molecule. A later candidate moleculewas compound 6 in reference 1. As published in Silverman 2003,“inactivation of GABA-AT was observed with 6, but when 2-mercaptoethanolwas added to the incubation mixture, no inactivation occurred.” The samepublication goes on to explain that the lack of activity in the presenceof 2-mercaptoethanol is an indication that the GABA-AT first acts oncompound 6 to form an alpha-beta unsaturated ketone(3-oxo-4-methylenyl-1-cyclopentanoic acid, compound 8 in thatpublication). The mercaptoethanol then reacts with the alpha-betaunsaturated ketone before it can inactivate the enzyme. This isundesirable because it indicates that the reactive intermediate escapesthe enzyme. To correct this deficiency, compound 6 was synthesized,which inactivated GABA-AT, even in the presence of 2-mercaptoethanol, soit was a true mechanism-based inactivator, and the reactive intermediatedoes not escape the enzyme prior to inactivation.

Once GABA-AT has been inactivated, it takes a number of days for thebrain to synthesize new GABA-AT to replace the inactivated enzyme.Information Petroff, Ognen A. C.; Rothman, Douglas L.; “Measuring HumanBrain GABA In Vivo, Effects of GABA-Transaminase Inhibition withVigabatrin”, Molecular Neurobiology, 1998, 16(1), 97-121 demonstratedthat brain GABA levels remain substantially elevated for several daysafter administration of a single dose of vigabatrin. This observation isconsistent with the theory that it takes several days for the brain torestore the GABA-AT activity.

Gabaergic drugs are also useful in treating Huntington's chorea ((a)Perry, T. L.; Hansen, S.; Lesk, D.; Kloster, M. “Amino Acids in Plasma,Cerebrospinal Fluid, and Brain of Patients with Huntington's Chorea.”Adv. Neurol. 1972, 1, 609. (b) McGeer, P. L.; McGeer, E. G. “The GABASystem and Function of the Basal Ganglia: Huntington's Disease.” InGABAin Nervous System Function Roberts, E.; Chase, T. N.; and Tower, D. B.;Eds.; Raven Press: New York, 1976; pp. 487-495. (a) Butterworth, J.;Yates, C. M.; Simpson, J. “Phosphate-activated glutaminase in relationto Huntington's disease and agonal state.” J. Neurochem. 1983, 41, 440.(b) Spokes, E. G. S. “Brain temperature after death.” Adv. Exp. Med.Biol. 1978, 123, 461. (c) Wu, J. Y.; Bird, E. D.; Chen, M. S.; Huang, W.M. “Abnormalities of neurotransmitter enzymes in Huntington's chorea.”Neurochem. Res. 1979, 4, 575. (d) Iversen, L. L.; Bird, E. D.; Mackay,A. V. P.; Rayner, C. N. “Analysis of glutamate decarboxylase inpost-mortem brain tissue in Huntington's chorea.” J. Psychiat. Res.1974, 11, 255., Parkinson's disease Nishino, N.; Fujiwara, H.;Noguchi-Kuno, S.-A.; Tanaka, C. “GABA receptor but not muscarinicreceptor density was decreased in the brain of patients with Parkinson'sdisease.” Jpn. J. Pharmacol. 1988, 48, 331. Maker, H. S.; Weiss, C.;Weissbarth, S.; Silides, D. J.; Whetsell, W. “Regional activities ofmetabolic enzymes and glutamate decarboxylase in human brain.” Ann.Neurol. 1981, 10, 377. (b) Rinne, U. K.; Laaksonen, H.; Riekkinen, P.;Sonninen, V. “Brain glutamic acid decarboxylase activity in Parkinson'sdisease.” Eur. Neurol. 1974, 12, 13. (c) McGeer, P. L.; McGeer, E. G.;Wada, J. A.; Jung, E. “Effects of globus pallidus lesions andParkinson's disease on brain glutamic acid decarboxylase.” Brain Res.1971, 32, 425., Alzheimer's disease (a) Aoyagi, T.; Wada, T.; Nagai, M.;Kojima, F.; Harada, S.; Takeuchi, T.; Takahashi, H.; Hirokawa, K.;Tsumita, T. “Increased g-aminobutyrate aminotransferase activity inbrain of patients with Alzheimer's disease.” Chem. Pharm. Bull. 1990,38, 1748-1749. (b) Davies, P. “Neurotransmitter-related enzymes insenile dementia of the Alzheimer type.” Brain Res. 1979, 171, 319. (c)Perry, E. K.; Gibson, P. H.; Blessed, G.; Perry, R. H.; Tomlinson, B. E.“Neurotransmitter enzyme abnormalities in senile dementia. Cholineacetyltransferase and glutamic acid decarboxylase activities in necripsybrain tissue.” J. Neurol. Sci. 1977, 34, 247. (d) Bowen, D. M.; White,P.; Flack, R. H. A.; Smith, C. B.; Davison, N. A. “Brain-decarboxylaseactivities as indices of pathological change in senile dementia.” Lancet1974, 1, 1247. (e) Kodama, K.; Kaitani, H.; Nanba, M.; Kondo, T.;Mikame, F.; Yoshida, H.; Sato, K.; Yanaihara, N. “Neurotransmitteranalogs in body fluids of patients with dementia.” Shinkei Kagaku 1981,20, 496, and tardive dyskinesia Gunne, L. M.; Haeggstroem, J. E.;Sjoequist, B. “Association with persistent neuroleptic-induceddyskinesia of regional changes in brain GABA synthesis.” Nature (London)1984, 309, 347.

Published United States patent application number 20040023952 A1, Ser.No. 10/311,821, entitled Enhanced Brain Function by GABA-ergicStimulation describes how GABA-ergic drugs are useful in treatingvariety of age-associated disorders of cortical decline in the elderly.These “age-associated” disorders of cortical decline extend on acontinuum from normal age-related senescence to severe dementiasassociated with Alzheimer's disease and Parkinson's disease in an agingpopulation. Published patent application 20040023952 A1 is expresslyincorporated herein by reference.

Also unexpected is the fact that(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid has anactivity more than 100 times that of vigabatrin in vitro. Usingmicro-positron emission tomography imaging techniques, that(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid completelyblocks cocaine-induced increases in synaptic dopamine in the nucleusaccumbens as well as the expression of cocaine-induced conditioned placepreference at a dose 100 times lower than that measured with vigabatrin.

Abbreviations: CPP, conditioned place preference; GABA, γ-aminobutyricacid; GABA-AT, γ-aminobutryic acid aminotransferase; μPET,micro-positron emission tomography; NAc, nucleus accumbens; VFD, visualfield defect

It is an object of the present invention to deliver a GABAaminotransferase inhibitor to a patient in need thereof while reducingvisual field defects.

It is an object of this invention to treat patients using anirreversible inhibitor of GABA aminotransferase.

It is an object of the present invention to suppress dopamine levelsbelow the level attainable by administration of vigabatrin.

It is an object of the present invention to treat cocaine addictionusing low doses of a GABA aminotransferase inhibitor.

It is an object of the present invention to treat cocaine addictionusing (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid.

DESCRIPTION OF THE FIGURES

FIG. 1 is the structure for(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid

FIG. 2 is a graph showing the effects of(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid alone and coadministered with GABA on oocytes expressing human ρ1 GABA_(C)receptors.

FIG. 3 is a graph comparing the time-dependent effects of(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid on dopaminerelease from nucleus accumbens (NAc) by cocaine administration to ratsversus vigabatrin and a saline control

FIG. 4 is a diagram illustrating a hypothetical explanation of why(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid is likely toreduce collateral damage to the retina and neurological structures.

FIG. 5 is a chart summarizing the results of visual field testing dataat 45 and 90 days

DETAILED DESCRIPTION OF THE INVENTION

The content of all references cited herein is expressly incorporated byreference.

Vigabatrin is known in the literature and has been approved for use intreating epilepsy and seizures and has been studied for treatment ofdrug addiction. U.S. Pat. Nos. 6,906,099; 6,890,951; 6,828,349;6,593,367; 6,541,520; 6,395,783; 6,323,239; and 6,057,368 which describeand/or claim the use of vigabatrin in the treatment of addiction fromcocaine, nicotine, methamphetamine, morphine, heroin, ethanol,phencyclidine, methylenedioxymethamphetamine, and/or PCP. It is believedthat compounds of the present invention will treat or prevent addictionof all of the following drugs: mu opiod receptor agonists including butnot limited to, 3-methylfentanyl, 3-methylthiofentanyl, Acetorphine,Acetyl methadol, Acetyl-alpha-methylfentanyl, Acetylhydrocodone,Alfentanil, Allylprodine, Alphaacetylmethadol, Alphameprodine,Alphamethadol, Alpha-methylfentanyl, Alpha-methylthiofentanyl,Benzethidine, Benzylmorphine, Beta-hydroxy-3-methylfentanyl,Beta-hydroxyfentanyl, Betameprodine, Betamethadol, Betaprodine,Betascetylmethadol, Bezitramide, Buprenorphine, Butorphanol,Carfentanil, Cocaine, Codeine, Cyprenorphine, Desomorphine,Dextromoramide, Diampromide, Diethylthiambutene, Difenoxin,Dihydrocodeine, Dihydroetorphine, Dihydromorphine, Dimenoxadol,Dimepheptanol, Dimethylthiambutene, Dioxaphetyl butyrate, Diphenoxylate,Dipipanone, Drotebanol, Ethanol, Ethylmethylthiambutene, Ethylmorphine,Etonitazine, Etorphine, Etoxeridine, Fentanil, Fentanyl, Furethidine,Heroin (diacetyl morphine), Hydrocodone, Hydromorphinol, Hydromorphone,Hydroxypethidine, Isomethadone, Ketobemidone, LAAM(levoalphaacetylmethadol), Levomethorphan, Levomoramide,Levophenacylmorphan, Levorphanol, Meperidine, Metazocine, Methadone,Methamphetamine, Methyldesorphine, Methyldihydromorphine, Metopon,Morpheridine, Morphine, MPPP (1-methyl-4-phenyl-4-propionoxypiperidine),Myrophine, Nalorphine, Nepetalactone, Nicocodeine, Nicomorphine,Nicotine; Noracymethadol, Norlevorphanol, Normethadone, Normorphine,Norpipanone, Opium, Oripavine, Oxycodone, Oxymorphone,Para-fluorofentanyl, Pentazocine, PEPAP(1-(2phenylethyl)-4-phenyl-acetoxypiperidine), Phenampromide,Phenazocine, Phenedoxone, Phenomorphan, Phenoperidine, Pholcodin,Piminodine, Piritramide, Proheptazine, Properidine, Propiram,Propxyphene, Racemethorphan, Racemoramide, Racemorphan, Remifentanil,Sufentanil, Tapentadol, Tapentadol, Thebaine, Thiofentanyl, Tilidine,Tramadol, Trimeperidine; dopamine reuptake inhibitors; CB1 receptoragonists, alpha adrenergic receptor agonists, dopamine receptoragonists; dopamine reuptake inhibitors, GABA agonists, nicotinicreceptor agonists. Addictive drugs to which the present invention isapplicable can be readily identified from 21 C.F.R. §1308 Schedules ofControlled Substances, et seq, which is expressly incorporated byreference.

It is well established that the neurochemical response to cocaine andother drugs of abuse is characterized by a rapid elevation in therelease of dopamine in the nucleus accumbens (NAc). Dewey, Stephen L.;Morgan, Alexander E.; Ashby, Charles R. Jr.; Horan Bryan; Kushner,Stephanie A.; Logan, Jean; Volkow, Nora D.; Fowler, Joanna S.; Gardner,Eliot L.; Brodie, Jonathan D.; A Novel Strategy for the Treatment ofCocaine Addiction. Synapse 1998, 30(2), 119-129 This increase indopamine, and associated behaviors, can be antagonized by an increase inthe concentration of γ-aminobutyric acid (GABA), which has been shown tooccur with use of the epilepsy drug vigabatrin (1, Sabril™), a knownmechanism-based inactivator. Silverman, R. B. Mechanism-Based EnzymeInactivation: Chemistry and Enzymology, Vols. I and II; CRC Press; BocaRaton, Fla.; 1988. (b) Silverman, R. B. Mechanism-Based EnzymeInactivators. Methods Enzymol. 1995, 249, 240-283 of γ-aminobutyric acidaminotransferase (GABA-AT). Lippert, B.; Metcalf, B. W.; Jung, M. J.;Casara, P.; 4-Aminohex-5-Enoic Acid, A Selective Catalytic Inhibitor of4-Aminobutyric Aminotransferase In Mammalian Brain. Eur. J. Biochem.1977, 74, 441-445. Vigabatrin is currently marketed for the treatment ofinfantile spasms (West's Syndrome) and refractory partial complexseizures in 65 countries, including the United States.

Vigabatrin also has been found to have utility in the treatment ofstimulant addiction. (Karila, L.; Gorelick, D.; Weinstein, A.; Noble,F.; Benyamina, A.; Coscas, S.; Blecha, L.; Lowenstein, W.; Martinot, J.L.; Reynaud, M.; Lepine, J. P. New treatments for cocaine dependence: afocused review. Internat. J. Neuropsychopharmacol. 2008, 11(3), 425-438.(b) Peng, X.-Q.; Li, X.; Gilbert, J. G.; Pak, A. C.; Ashby, C. R.;Brodie, J. D.; Dewey, S. L.; Gardner, E. L.; Xi, Z.-X. Gamma-vinyl GABAinhibits cocaine-triggered reinstatement of drug-seeking behavior inrats by a non-dopaminergic mechanism. Drug Alcohol Depend. 2008, 97(3),216-225. Vigabatrin has been specifically shown to be effective inanimal models for cocaine, (a) Perry, T. L.; Hansen, S.; Lesk, D.;Kloster, M. “Amino Acids in Plasma, Cerebrospinal Fluid, and Brain ofPatients with Huntington's Chorea.” Adv. Neurol. 1972, 1, 609. (b)McGeer, P. L.; McGeer, E. G. “The GABA System and Function of the BasalGanglia: Huntington's Disease.” In GABA in Nervous System FunctionRoberts, E.; Chase, T. N.; and Tower, D. B.; Eds.; Raven Press: NewYork, 1976; pp. 487-495) nicotine, (Dewey, Stephen L.; Brodie, JonathanD.; Gersimov, Madina; Horan, Bryan; Gardner, Eliot L.; Ashby, Charles R.Jr.; A Pharmaceutical Strategy for the Treatment of Nicotine Addiction.Synapse 1999, 31(1), 76-86), methamphetamine, heroin, ethanol (Gersimov,Madina R.; Ashby, Charles R. Jr.; Gardner, Eliot L.; Mills, Mark J.;Brodie, Jonathan D.; Dewey, Stephen L.; Gamma-vinyl GABA InhibitsMethamphetamine, Heroin, or Ethanol-Induced Increases in NucleusAccumbens Dopamine. Synapse 1999, 34(1), 11-19.), and combinationaddictions (Stromberg, Michael F.; Mackler, Scott A.; Volpicelli, JosephR.; O'Brien, Charles P.; Dewey, Stephen L.; The effect ofgamma-vinyl-GABA on the consumption of concurrently available oralcocaine and ethanol in the rat. Pharmacol. Biochem. Behav. 2001, 68,291-299). Vigabatrin treatment also is effective for stimulant addictionin humans ((a) Brodie, Jonathan D.; Figueroa, Emilia; Dewey, Stephen L.;Treating Cocaine Addiction: From Preclinical to Clinical TrialExperience with γ-vinyl GABA. Synapse 2003, 50(3), 261-265. (b) Brodie,Jonathan D.; Figueroa, Emilia; Laska, Eugene M.; Dewey, Stephen L.;Safety and Efficacy of γ-Vinyl GABA (GVG) for the Treatment ofMethamphetamine and/or Cocaine Addiction. Synapse 2005, 55(2),122-125.), including a recently reported randomized, double-blind,placebo-controlled, trial of 103 subjects (Brodie, Jonathan D.; Case,Brady G.; Figueroa, Emilia; Dewey, Stephen L.; Robinson, James A.;Wanderling, Joseph A.; Laska, Eugene M.; Randomized, Double-Blind,Placebo-Controlled Trial of Vigabatrin for the Treatment of CocaineDependence in Mexican Parolees. Am. J. Psychiatry 2009, 166, 1269-12770,in which 28.0% of subjects treated with vigabatrin achieved abstinencecompared to 7.5% of subjects treated with placebo.

The acceptance of vigabatrin for the treatment of both epilepsy and as apotential treatment for stimulant addiction has been hampered primarilyby concerns about abnormalities of the peripheral visual field (visualfield defects or visual field defect) in 25-50% of patients followingchronic administration of vigabatrin. Willmore, L. James; Abelson, MarkB.; Ben-Menachem, Elinor; Pellock, John M.; Shields, Donald; Vigabatrin:2008 Update. Epilepsia 2009, 50(2), 163-173; Wild, John M.; Chiron,Catherine; Ahn, Hyosook; Baulac, Michel; Bursztyn, Joseph; Gandolfo,Enrico; Goldberg, Ivan; Goni, Francisco Javier; Mercier, Florence;Nordmann, Jean-Philippe; Safran, Avinoam B.; Schiefer, Ulrich; Perucca,Emilio; Visual Field Loss in Patients with Refractory Partial EpilepsyTreated with Vigabatrin. CNS Drugs 2009, 23(11), 965-982 Shorterduration exposure in connection with studies of the treatment ofstimulant addiction with vigabatrin does not show any occurrence ofvisual field defect, which corroborates the prevailing belief that thedevelopment of visual field defect results from prolonged exposure tovigabatrin. Fechtner, Robert D.; Khouri, Albert S.; Figueroa, Emilia;Ramirez, Marina; Federico, Martha; Dewey, Stephen L.; Brodie, JonathanD.; Short-term Treatment of Cocaine and/or Methamphetamine Abuse withVigabatrin-Ocular Safety Pilot Results. Arch. Ophthalmol. 2006, 124,1257-1262 Treatment of addictive disorders is usually long term orchronic therapy. The long term administration of vigabatrin is knowncauses visual field defects. The mechanism leading to the visual fielddefect is not known, but it remains an active area of research. Visualfield defects might occur from elevated GABA levels as a result ofinactivation of GABA-AT, could be a direct toxic effect of vigabatrin,could be the consequence of an enzymatically produced byproduct from oneof the enzyme inactivation mechanisms, or some combination of thesepotential mechanisms.

A new synthetic compound,(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid (2), wasdesigned as a mechanism-based inactivator of GABA-AT, which couldgenerate a more reactive intermediate along the pathway to attachment tothe active site of GABA-AT via a Michael addition Pan, Y.; Qiu, J.;Silverman, R. B.; Design, Synthesis and biological Activity for aDifluoro-substituted, conformationally-rigid Vigabatrin Analogue As aPotent γ-Aminobutyric Acid Aminotransferase Inhibitor. J. Med. Chem.2003, 46, 5292-5293. In contrast to the high K₁ value (3.2 mM¹²; 10 mM³)reported for vigabatrin as an inactivator of GABA-AT, the new syntheticGABA-AT inactivator (2) has a K₁ value of 31 μM¹². A comparison of thek_(inact)/K₁ values (a measure of the efficiency of the inactivator)indicated that (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoicacid is 186 times more effective as an inactivator of GABA-AT thanvigabatrin under suboptimal conditions (at optimal conditions forsubstrate turnover, the rate of inactivation is too rapid to measure;these values were obtained at a pH and temperature well below theoptimum). Despite irreversibility of the inhibition, the low potency ofvigabatrin translates into treatment doses of 1-3 g/day (U.S. Labelingfor Sabril®http://www.lundbeckinc.com/USA/products/CNS/Sabril/sabril_PI_CPS.pdf.Because (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic aciddisplayed superior enzyme inactivation properties compared tovigabatrin, we have carried out further pharmacological studies with(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid. The affinityof (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid atGABA_(A) and GABA_(B) receptors and its activity at the GABA_(C)receptor as well as at four GABA transporter subtypes expressed eitherendogenously in neurons and astrocytes or recombinantly in mammaliancell lines was determined. Because of the preponderance of dataindicating vigabatrin is effective for the treatment of addiction, thepreviously reported¹¹ lack of visual field defect observed for shortvigabatrin exposure durations required for the treatment of stimulantaddiction, and the relatively short duration of drug exposure needed foraddiction treatment, the effect of(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid oncocaine-induced conditioned place preference in rats (an animal modelfor effectiveness of addiction treatments) also was investigated.Mechanistic similarities between vigabatrin and(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid for thetreatment of addiction were also investigated by μPET imaging to measurethe ability of (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoicacid to antagonize cocaine-induced increases in synaptic nucleusaccumbens dopamine.

While the compound (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoicacid has been shown to be a GABA aminotransferase inhibitor likevigabatrin, surprisingly(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid does notinhibit the reuptake of GABA.

Materials

Vigabatrin, (R)-baclofen, GABA, isoguvacine, sodium pyruvate,theophylline, gentamycin, and all buffer reagents were purchased fromSigma-Aldrich (St. Louis, Mo., USA).(1S,3S)-3-Amino-4-difluoromethylenyl-1-cyclopentanoic acid (2) wassynthesized as reported previously.¹² [³H]GABA (35 or 40.0 Ci/mmol) and[³H]muscimol (36.6 Ci/mmol) were purchased from PerkinElmer (Boston,Mass., USA). All reagents for cell culturing were purchased fromInvitrogen (Paisley, UK). Cocaine USP was provided by the NationalInstitute on Drug Abuse (NIDA). All animals were adult maleSprague-Dawley rats (200-225 g, supplied by Taconic Farms, Germantown,N.Y.).

GABA Uptake Assay

[³H]GABA Uptake Assay at Human GABA Transporters

tsA201 cells were cultured in GlutaMAX-I DMEM supplemented with 10%fetal bovine serum, penicillin (100 U/ml), and streptomycin (100 μg/ml)at 37° C. in a humidified atmosphere of 95% air and 5% CO₂. The plasmidsencoding hGAT-1, hBGT-1, hGAT-2, and hGAT-3, (Kvist, T.; Christiansen,B.; Jensen, A. A.; Bräuner-Osborne, H. The four human gamma aminobutyricacid (GABA) transporters: pharmacological characterization andvalidation of a highly efficient screening assay. Comb. Chem. HighThroughput Screen 2009, 12, 241-249) respectively, were transfected intotsA201 cells using PolyFect according to the protocol of themanufacturer (Qiagen, West Sussex, UK). The next day, the tsA201 cellstransiently expressing each of the four human GABA transporter subtypeswere split into poly-D-lysine-coated white 96-well plates (PerkinElmer).The pharmacological assays were performed 36-48 h after transfectionexactly as described previously Christiansen, B.; Meinild, A. K.;Jensen, A. A; Bräuner-Osborne, H. Cloning and characterization of afunctional human gamma-aminobutyric acid (GABA) transporter, humanGAT-2. J. Biol. Chem. 2007, 282, 19331-19341. In brief, assay buffersupplemented with 30 nM [³H]GABA and test compounds was added to thecells, and the uptake of [³H]GABA was determined after incubation at 37°C. for 3 min. Quantification was performed by using Microscint™20scintillation fluid (PerkinElmer) and a Packard TopCount microplatescintillation counter.

[³H]GABA Uptake Assay at Mouse GABA Transporters

Cortical astrocytes were cultured essentially as previously described.Hertz L, Juurlink B H J, Hertz E, Fosmark H and Schousboe A. Preparationof Primary Cultures of Mouse (Rat) Astrocytes, in A Dissection andTissue Culture Manual of the Nervous System (Shahar A, de Vellis J,Vernadakis A and Haber B eds) pp 105-108, Alan R. Liss, Inc., New York,1989 The neopallium was removed from new born NMRI mice (Taconic,Denmark) and passed through an 80 μm nylon sieve and cultured inmodified Dulbecco's modified Eagle's medium with fetal calf serum. Thecalf serum was lowered from 20% to 10% over three weeks, and finally theastrocytes were allowed to differentiate using 0.25 mM dibutyryl cyclicAMP during the last week of growth.

Cortical neurons were cultured essentially as previously described byremoving the neopallium of 15-day old NMRI embryos by dissectionfollowed by mild trypsination. Hertz E, Yu A C H, Hertz L, Juurlink B HJ and Schousboe A. Preparation of Primary Cultures of Mouse CorticalNeurons, in A Dissection and Tissue Culture Manual of the Nervous System(Shahar A, de Vellis J, Vernadakis A and Haber B eds) pp 183-186, AlanR. Liss, Inc., New York, 198. The neurons were cultured in 10% fetalcalf serum and, after 48 h, cytosine arabinoside was added to a finalconcentration of 20 μM to prevent glial proliferation. Four cultures ofstably transfected Human Embryonic Kidney (HEK)-293 cells expressingmGAT1-4 were prepared by the method previously reported. White H S,Sarup A, Bolvig T, Kristensen A S, Petersen G, Nelson N, Pickering D S,Larsson O M, Frolund B, Krogsgaard-Larsen P and Schousboe A. CorrelationBetween Anticonvulsant Activity and Inhibitory Action on GlialGamma-Aminobutyric Acid Uptake of the Highly Selective MouseGamma-Aminobutyric Acid Transporter 1 Inhibitor3-Hydroxy-4-Amino-4,5,6,7-Tetrahydro-1,2-Benzisoxazole and ItsN-Alkylated Analogs. J Pharmacol Exp Ther. 2002, 302, 636-644 The stablecell lines are under the selection pressure of blasticidin-S at 5 μg/mL.Determinations of the IC₅₀ values were conducted as described earlier.Bolvig T, Larsson 0 M, Pickering D S, Nelson N, Falch E,Krogsgaard-Larsen P and Schousboe A. Action of Bicyclic Isoxazole GABAAnalogues on GABA Transporters and Its Relation to AnticonvulsantActivity. Eur. J. Pharmacol. 1999, 375, 367-374 In brief, [³H]GABAuptake was assessed at 37° C. for 3 min on desired cells in PBS buffercontaining 1 μM GABA, 13 nM [³H]GABA, and test compound. Radioactivitywas measured using Microscint™20 scintillation fluid (PerkinElmer) and aPackard TopCount microplate scintillation counter.

GABA Receptor Binding Assays Receptor Preparations

GABA_(A) and GABA_(B) binding assays were performed using rat brainsynaptic membranes of cortex and the central hemispheres from adult maleSprague-Dawley rats with tissue preparation as earlier described.Ransom, R. W.; Stec, N. L. Cooperative modulation of [³H]MK-801 bindingto the N-methyl-D-aspartate receptor-ion channel complex by L-glutamate,glycine, and polyamines. J. Neurochem. 1988, 51, 830-836. On the day ofthe assay, the membrane preparation was quickly thawed, suspended in 40volumes of ice-cold 50 mM Tris-HCl buffer (pH 7.4) using an UltraTurraxhomogenizer and centrifuged at 48,000 g for 10 min at 4° C. This washingstep was repeated four times. The final pellet was resuspended inincubation buffer and the binding assay carried out as detailed below.

GABA_(A) Receptor Activity Assay

Rat brain synaptic membranes (100 μg protein/aliquot) prepared above inTris-HCl buffer (50 mM, pH 7.4) were incubated with [³H]muscimol (5 nM)and 100 μM of compound 2 at 0° C. for 60 min in a total volume of 250μl. GABA (1 mM) was used to define non-specific binding. The bindingreaction was terminated by rapid filtration through GF/B unifilters(PerkinElmer) using a 96-well Packard FilterMate cell harvester,followed by washing with 3×250 μl of ice-cold binding buffer, drying,and adding scintillation fluid, as described for the [³H]GABA uptakeassay.

GABA_(B) Receptor Binding Assay

For [³H]GABA binding to the GABA_(B) receptors, rat brain synapticmembranes (200 μg protein/aliquot) were suspended in Tris-HCl buffer (50mM+2.5 mM CaCl₂, pH 7.4) and incubated with [³H]GABA (5 nM), isoguvacine(40 μM), and 100 μM of compound 2 at 25° C. for 45 min in 1 ml totalvolume. Isoguvacine serves to saturate GABA_(A) receptors. Hill, D. R.;Bowery, N. G. ³H-baclofen and ³H-GABA bind to bicuculline-insensitiveGABA_(B) sites in rat brain. Nature 1981, 290, 149-152 Non-specificbinding was determined using 100 μM (R)-baclofen. Binding was terminatedby filtration through Whatman GF/C filters, using a Brandell M-48R CellHarvester; filters were washed with 3×3 ml of ice-cold buffer, andfilter-bound radioactivity was counted in a Packard Tricarb 2100 liquidscintillation analyzer using 3 ml of Opti-fluor scintillation fluid(PerkinElmer).

Electrophysiology

Expression of ρ1 in Xenopus leavis oocytes

Human ρ1 cDNA encapsulated in pcDNA1.1 was linearized with Notl.Linearized cDNA was transcribed to mRNA using the T7 “mMESSAGE mMACHINE”kit (Ambion Inc. Austin, Tex., USA) as previously described. Chebib, M;Duke, R. K.; Allan, R. A.; Johnston, G. A. R. The effects ofcyclopentane and cyclopentene analogs of GABA at recombinant GABA_(C)receptors. Eur. J. Pharmacol. 2001, 430, 185-192. GABA_(C) receptoractivity assays were performed in oocytes harvested from Xenopus laevis(housed in the Department of Veterinary Science at the University ofSydney) and defolliculated. The oocytes were stored in ND96 solution (inmM) NaCl (96), KCl (2), MgCl₂ (1), CaCl₂ (1.8), HEPES (hemi-Na salt; 5)supplemented with sodium pyruvate (2.5), theophylline (0.5), and 50μg/ml⁻¹ gentamycin for 2-5 days post-injection.

GABA_(C) receptor electrophysiological assay

Electrophysiological methods were performed as previously described.Hertz L, Juurlink B H J, Hertz E, Fosmark H and Schousboe A. Preparationof Primary Cultures of Mouse (Rat) Astrocytes, in A Dissection andTissue Culture Manual of the Nervous System (Shahar A, de Vellis J,Vernadakis A and Haber B eds) pp 105-108, Alan R. Liss, Inc., New York,1989 Stage V-VI oocytes were injected with 10 ng 50 nl⁻¹ of ρ1 mRNA andthen stored at 16° C. Recordings of receptor activity were obtained for2-5 days by a two-electrode voltage clamp by means of a Geneclamp 500amplifier (Axon Instruments Inc., Foster City, Calif.), a MacLab 2erecorder (AD Instruments, Sydney, NSW), and Chart version 3.6.3 program.Oocytes were voltage clamped at −60 mV, and the preparation wascontinually perfused with ND96 solution at room temperature. Compound 2(CPP-115) (100 μM) dissolved in ND96 was applied in the absence andpresence of GABA, respectively, until maximum current was reached, atwhich time the oocytes were washed for 5 to 10 min to allow completerecovery of response to GABA (1 μM). Compound 2 (CPP-115) was tested onthree oocytes from at least two harvests.

Cocaine-Induced Conditioned Place Preference (CPP)

A non-biased approach was used for all CPP studies. Specifically,animals were pretested in the CPP chambers for a pre-existing chamberbias. Any animals that spent more than 70% of their time in any chamberwere eliminated from the study. Thus, only animals that demonstrated nopre-existing chamber bias were used in the study.

In all rodent studies (n=8/group) animals were allowed to acclimate tothe animal housing facility for at least 5 days prior to beginning theexperiments. CPP chambers were used as previously described, (Ashby, C.R., Jr.; Paul, M.; Gardner, E. L.; Gerasimov, M. R.; Dewey, S. L.;Lennon, I. C.; Taylor, S. J. C. Systemic administration of1R,4S-4-amino-cyclopent-2-ene-carboxylic acid, a reversible inhibitor ofGABA transaminase, blocks expression of conditioned place preference tococaine and nicotine in rats. Synapse (New York, N.Y., United States)2002, 44(2), 61-63.) except instead of one chamber being entirely whiteand the other black, one chamber was entirely light blue with astainless steel floor, and the second chamber was light blue withhorizontal black stripes (2.5 cm wide) spaced 3.8 cm apart with a smoothPlexiglass floor. In all CPP studies with 2, the saline volume was (1ml/kg), the cocaine doses were 20 mg/kg, and the dosage of 2 was 1.0mg/kg. The saline, cocaine, and Compound 2 (CPP-115) were all injectedintraperitoneally (i.p.). The conditioning procedure for the acquisitionphase consisted of 12 sessions carried out consecutively over 12 days.The CPP pairings were: 1) saline/saline; 2) saline/cocaine; 3) Compound2 (CPP-115)/saline, and 4) saline/cocaine+Compound 2 (CPP-115). Animalsin each group were randomly assigned to a 2×2 factorial design with onefactor being the pairing chamber and the other factor being the order ofconditioning.

Animals that received either saline or cocaine were injected andconfined to the appropriate compartment for 30 min.(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid injectionswere given 2.5 h prior to saline or cocaine injections. This was done asit has been shown that GABA levels reach maximal values 3 to 4 hfollowing the administration of(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid. On the testday (day 12) neither drugs nor saline was administered, and the animalswere allowed to move freely between both chambers for 15 min. The amountof time spent in each chamber was recorded using an automated infraredbeam electronically coupled to a timer. For the expression phase of CPPto cocaine, the animals were habituated and conditioned to cocaine asdescribed in the acquisition studies, but no animals in the expressionstudies were given 2 on conditioning days. On the test day, the animalsbeing tested received either saline or Compound 2 (CPP-115) 2.5 h priorto their being placed in the apparatus and allowed free access to bothchambers for 15 min. A time period of 2.5 hours was selected becauseprevious studies demonstrated that this was the optimal pretreatmentinterval allowing for a maximal increase in GABA concentrations. Dewey,S. L.; Morgan, A. E.; Ashby Jr., C. R.; Horan, B.; Kushner, S. A.;Logan, J.; Volkow, N. D.; Fowler, J. S.; Gardner, E. L.; Brodie, J. D. Anovel strategy for the treatment of cocaine addiction. Synapse 1998, 30,119-129.

μPET Imaging Studies

Using separate adult animals (male Sprague-Dawley rats, n=2) μPETstudies were performed using a Concorde Microsystems R4. Baseline¹¹C-raclopride binding was examined in anesthesized (ketamine/xylazine)animals. ¹¹C-raclopride (20.4 min half-life) is selective for thedopamine family of receptors and competes directly with dopamine forreceptor binding. Thus, drug-induced increases in brain dopamine producea decrease in ¹¹C-raclopride binding while dopamine depletion producesan increase in binding. Approximately 2 h following these baselinescans, animals received an intravenous injection of cocaine (5 mg/kg)followed 5 min later by a second injection of ¹¹C-raclopride.Approximately 2 h following this scanning session, animals receivedcompound (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid (0.5mg/kg). Approximately 2.5 h following the administration of(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid, animalsreceived a second intravenous dose of cocaine (5.0 mg/kg) followed 5 minlater with a third injection of ¹¹C-raclopride.

Effects of (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid(2) at GABA transporters and receptors Interaction of(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid with GABAtransporters

(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid displayed noinhibitory activity at 1 mM concentration at GABA transporters inneurons, astrocytes, or mammalian cells recombinantly expressing humanor mouse transporter subtypes (Table 1). The pharmacological propertiesof (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid werecharacterized in tsA201 or HEK293 cells transiently expressing the fourhuman or mouse GABA transporter subtypes (human hGAT-1, hBGT-1, hGAT-2,hGAT-3, and mouse mGAT1-4, respectively).

TABLE 1 Effects of GABA and Compound 2 (CPP-115) on GABA uptake inneurons, astrocytes, and human and mouse GABA transporter-expressingcells (1S,3S)-3-amino-4- difluoromethylenyl- 1-cyclopentanoic GABA acidIC₅₀ (μM) IC₅₀ (μM) hGAT-1 uptake >1000 10^(a) hBGT-1 uptake >100026^(a) hGAT-2 uptake >1000 11^(a) hGAT-3 uptake >1000 10^(a) mGAT1uptake >1000 17^(b) mGAT2 uptake >1000 51^(b) mGAT3 uptake >1000 15^(b)mGAT4 uptake >1000 17^(b) neuron uptake >1000  8^(b) astrocyteuptake >1000 32^(b) ^(a)data from Kvist, T.; Christiansen, B.; Jensen,A. A.; Bräuner-Osborne, H. The four human gamma aminobutyric acid (GABA)transporters: pharmacological characterization and validation of ahighly efficient screening assay.Comb. Chem. High Throughput Screen2009, 12, 241-249 ^(b)data from Bolvig T, Larsson O M, Pickering D S,Nelson N, Falch E, Krogsgaard-Larsen P and Schousboe A. Action ofBicyclic Isoxazole GABA Analogues on GABA Transporters and Its Relationto Anticonvulsant Activity. Eur. J. Pharmacol. 1999, 375, 367-374.

To investigate a possible interaction of(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid with GABAreceptors, the compound was tested for its ability to displace [³H]GABAbinding to ionotropic GABA_(A) receptors or metabotropic GABA_(B)receptors in rat brain cortical homogenate. At a concentration of 100μM, no inhibition of binding was observed at either receptor tested,whereas 1 mM cold GABA inhibited radioligand binding as expected (Table2). Furthermore, (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoicacid was tested for activity at recombinant human ρ1 GABA_(C) receptorsexpressed in oocytes and was found to exhibit no effect as an agonist orantagonist at a concentration of 100 μM (FIG. 1).

TABLE 2 Effect of (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoicacid on GABA_(A) and GABA_(B) receptors evaluated in binding assays IC₅₀(μM) (1S,3S)-3-amino-4- difluoromethylenyl- 1-cyclopentanoic acid GABA[³H]muscimol competition >100 0.049^(a) (GABA_(A) receptor) [³H]GABAcompetition^(b) >100 0.013^(a) (GABA_(B) receptor) ^(a)data fromWellendorph, P.; Høg, S.; Greenwood, J. R.; de Lichtenberg, A.; Nielsen,B.; Frølund, B.; Brehm, L.; Clausen, R. P.; Bräuner-Osborne, H. Novelcyclic gamma-hydroxybutyrate (GHB) analogs with high affinity andstereoselectivity of binding to GHB sites in rat brain. J. Pharmacol.Exp. Ther. 2005, 315, 346-351. ^(b)a high concentration of isoguvacinewas added to ensure saturation of GABA_(A) receptor sites

Cocaine-Induced Conditioned Place Preference (CPP) Studies

Effect of compound (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoicacid on dopamine release from nucleus accumbens (NAc) by cocaineadministration to rats.

To compare the pharmacological effects of(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid to thepreviously reported effect of vigabatrin, the effect of cocaineadministration on NAc-released dopamine was determined. In thesepreliminary μPET imaging studies, cocaine reduced ¹¹C-raclopride bindingby an average of 22%, consistent with an increase in synaptic dopamine.However, when treated with(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid, there was noeffect of cocaine on ¹¹C-raclopride binding. That is, ¹¹C-raclopridebinding was similar to the control data, consistent with(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid (0.5 mg/kg)producing a complete blockade of cocaine-induced increases in synapticdopamine at a dose 600 times lower than the 300 mg/kg dose of vigabatrinthat was effective previously. Ashby, C. R., Jr.; Rohatgi, R. i;Ngosuwan, J.; Borda, T.; Gerasimov, M. R.; Morgan, A. E.; Kushner, S.;Brodie, J. D.; Dewey, S. L. Implication of the GABA_(B) receptor ingamma vinyl-GABA's inhibition of cocaine-induced increases in nucleusaccumbens dopamine. Synapse (New York) 1999, 31(2), 151-153.

Effect of (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid onthe expression of CPP

Increases in NAc dopamine following administration of cocaine produces adose-dependent and profound effect on the expression of CPP in rats. CPPis a well-documented model that assesses the saliency of drugs of abusein a drug-free state. The ability to pharmacologically block theexpression of a cocaine-induced CPP suggests that these compounds mighthave an indication for treating cocaine addiction.

Cocaine produced a dose-dependent CPP response, with the most reliableand robust response occurring at 20 mg/kg. Therefore, we chose a 20mg/kg cocaine dose with which to examine the effect of theadministration of (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoicacid on the expression of a cocaine-induced CPP. The results clearlyindicate that 1.0 mg/kg of 2 blocked the expression of cocaine-inducedCPP. By itself, (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoicacid produced neither a CPP nor a conditioned aversive response,indicating that 2 exhibits no abuse potential. These data areinteresting in that similar findings with vigabatrin required a dose of300 mg/kg, while the effects of(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid were obtainedusing a dose of only 1.0 mg/kg. Specifically, in the saline/salinepairings, animals spent an equal amount of time in both chambers(7.2±2.2 versus 7.8±2.9 min). However, in the saline/cocaine pairings,animals spent a significantly greater amount of time in thecocaine-paired chamber 12.2±1.7 versus 4.8±2.8 min (p<0.01, Student'stwo-tailed t-test). In the saline/Compound 2 (CPP-115) pairings, animalsspent an equal amount of time in both chambers (8.1±3.2 versus 6.9±3.9min), suggesting that(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid did notproduce a CPP on its own. In thecocaine/saline+(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoicacid pairings, animals again spent an equal amount of time in bothchambers (7.9±1.5 versus 7.1±1.9 min), demonstrating that at a dose of1.0 mg/kg, (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acidcompletely blocked the expression of a cocaine-induced CPP, which is 300times the effect of vigabatrin.

Because of the importance of GABAergic effects on a variety ofneurological disorders and the inherent complexity of this systemresulting from multiple subtypes of receptors and transporters, it iscrucial that potential therapeutic compounds are selective for specificcomponents of the GABAergic system. In this study we evaluated theselectivity profile of a recently described GABA-AT inhibitor,(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid (2). We findthat (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid does notaffect GABA uptake in recombinantly expressed human and mouse GABAtransporters or in mouse cortical astrocytes or neurons. Furthermore,(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid displays noaffinity for GABA_(A) or GABA_(B) receptors and is neither an agonistnor an antagonist for GABA_(C) receptors. Although(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid was nottested for functional activity at GABA_(A) and GABA_(B) receptors, thusnot ruling out a possible allosteric mechanism, the structuralresemblance of (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoicacid to GABA justifies ruling out binding to the GABA site overallosteric site. As previously reported, the principal GABAergic site ofaction of (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acidappears to be GABA-AT, the enzyme that catabolizes GABA. Sherif, F. M.;Ahmed, S. S. Basic aspects of GABA transaminase in neuropsychiatricdisorders. Clin. Biochem. 1995, 28(2), 145-54. Because of theeffectiveness of vigabatrin, an irreversible inactivator of GABA-AT, onthe reversal of specific addiction-associated biochemical and behavioralmeasures to a variety of drugs of abuse,(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid wasinvestigated for its ability to block cocaine-induced increases in NAcdopamine concentrations by μPET in sedated animals, an indicator ofaddictive behavior. Further, we extended these biochemical findings to abehavioral measure, the expression of a cocaine-induced CPP.

It is likely that the smaller increase in NAc dopamine levels producedby an acute cocaine challenge following either pretreatment withvigabatrin or (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acidis what underlies the effects we observed in both the μPET imaging andthe CPP studies. In fact, we observed that a dose of(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid that is 1/300(1.0 mg/kg) to 1/600 (0.5 mg/kg) that of vigabatrin (300 mg/kg)completely reversed cocaine-induced increases in synaptic dopamine aswell as in the expression of a cocaine-induced CPP. Given theeffectiveness and visual safety of vigabatrin in clinical trials for thetreatment of cocaine and/or methamphetamine addiction, in combinationwith its pre-clinical efficacy for the treatment of nicotine,methamphetamine, heroin, and ethanol abuse, it is likely that(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid also will beeffective in treating these addictive behaviors in humans. The potentialadvantage of (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid,however, is its much greater potency relative to vigabatrin, which couldmarkedly reduce its daily dosage relative to that of vigabatrin (1-3g/day). Furthermore, given the visual adverse effects of vigabatrinfrequently reported after long-term use in relation to its epilepsyindication, (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acidhas been evaluated for visual side effects to possibly provide analternative to vigabatrin for epilepsy patients internationally. If thepredominant cause of visual field defect in vigabatrin therapy is notdue to elevated GABA, as has been argued by some researchers, (Sills, G.J.; Butler, E.; Forrest, G.; Ratnaraj, N.; Patsalos, P. N.; Brodie, M.J. Vigabatrin, but not gabapentin or topiramate, producesconcentration-related effects on enzymes and intermediates of the GABAshunt in rat brain and retina. Epilepsia 2003, 44(7), 886-892) themechanistic differences between vigabatrin and(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid with regardto their inactivation of GABA-AT could contribute to less visual fielddefect in the case of(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid.

FIG. 2 shows the effects of GABA and CPP 115 on GABAc. GABA (1 μM; EC₈₀)(duration indicated by black bar) activated an inward current in oocytesexpressing human ρ1 GABA_(C) receptors clamped at −60 mV. CPP-115((1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid) (100 μM,duration indicated by the white bar) did not activate a current. Whenco-applied with GABA (1 μM), CPP 115 (100 μM) did not significantlyreduce the GABA response (p>0.05; n=3; Student t-test).

FIG. 3 shows the effect of the administration of a single dose of(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid, vigabatrinor a saline control on a cocaine induced dopamine surge prior to thecocaine challenge. Each drug was administered near their maximallyeffective therapeutic dose (1 mg/kg and 300 mg/kg), respectively. Thelevels of dopamine were measured by observing the displacement of¹¹C-raclopride from dopamine receptors by the presence of intersynapticdopamine using positron emission tomography in Sprague Dawley rats.(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid resulted in asurprising and significantly lower dopamine surge than is achievablewith vigabatrin under the same experimental conditions. Dopamine releasein the saline control group demonstrated a dopamine increase more than500 percent of basline. Those animals treated with vigabatrin showed adopamine increase more that 300 percent of baseline whereas the(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid groupdisplayed levels less than 300 times baseline. These data show that(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid is superiorto vigabatrin in its ability to reduce a cocaine induced dopamine surgeat the maximally effective therapeutic dose of each drug. Such areduction in the dopamine surge is expected to be more effective intreating cocaine addiction than vigabatrin at doses where GABA-AT ismaximally inactivated.

Treatment of Pain and Nervous System Disorders.

Gabaergic drugs are those which improve secretion or transmission ofGABA. These drugs as a family have been used to treat a wide variety ofnervous system disorders including Fibromyalgia, neuropathy, migrainesrelated to epilepsy, restless leg syndrome, and post traumatic distressdisorder. Gabaergic drugs include GABA_(A) and GABA_(B) receptorligands, GABA reuptake inhibitors, GABA aminotransferase inhibitors,GABA analogs, or molecules containing GABA itself. Preferred GABAergicdrugs include valproate and its derivatives, vigabatrin, pregabalin,gabapentin and tiagabine.

It is proposed that(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid may be usedto treat epilepsy, infantile spasms fibromyalgia, neuropathy, migrainesrelated to epilepsy, restless leg syndrome, and post traumatic distressdisorder at significantly lower doses that currently approved Gabaergicdrugs.

Published United States patent application number 20040023952 A1, Ser.No. 10/311,821, entitled Enhanced Brain Function by GABA-ergicStimulation describes how gabaergic drugs are useful in treating varietyof age-associated disorders of cortical decline in the elderly. These“age-associated” disorders of cortical decline extend on a continuumfrom normal age-related senescence to severe dementias associated withAlzheimer's disease and Parkinson's disease in an aging population.Published patent application 20040023952 A1 is expressly incorporatedherein by reference.

U.S. Pat. No. 6,939,876 describes and/or claims the use of vigabatrin inthe treatment to prevent addiction to opioid analgesics by coadministration of vigabatrin. The contents of U.S. Pat. No. 6,939,876 isexpressly incorporated herein by reference. It is believed that(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid will preventaddiction to opioid analgesics by administering the(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid to thepatient before, with or after administration of the opioid.

GABA aminotransferase inhibitors have been shown to be effective fortreatment of obsessive compulsive disorders including general anxietydisorder, pathological or compulsive gambling disorder, compulsiveeating (obesity), body dysmorphic disorder, hypochondriasis, pathologicgrooming conditions, kleptomania, pyromania, attention deficithyperactivity disorder and impulse control disorders in U.S. Pat. No.6,462,084, which is expressly incorporated by reference. Because(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid is a GABAaminotransferase inhibitor, it is anticipated it will have the sameactivity but without the visual field defects and at doses between 1/100to about 1/700 the dose of vigabatrin.

Dosages of (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acidare anticipated to be 2 to 2.5 mg/kg/day for treatment of epilepsy orabout 140-175 mg/day for an average 70 kg adult. The dose for treatmentof addiction is expected be to be only 0.05 to 0.2 mg/kg/day, or about3.0-15 mg/day for the average 70 kg adult.

The mechanism of inactivation of GABA is known. Nanavati, Shrenik M.,Silverman, Richard B. Mechanisms of inactivation of gamma-aminobutyricacid aminotransferase by the antiepilepsy drug gaba vinyl GABA(vigabatrin). J. Am. Chem. Society, 1991 113(24), 9341-9349; Pan, Yue,et al. Design, Synthesis and Biological Activity of aDiFluoro-Substituted Conformationally Rigid Vigabatrin Analogue as aPotent Aminobutyric Acid Aminotransferase Inhibitor. J. Med. Chem. 2003,46(25) 5292-5293).

Without wishing to be limited in theory, applicants believe that(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid will notcause or will significantly reduce visual defect damage as compared tovigabatrin at equally efficacious doses of both drugs. Referring to FIG.4, which shows a diagram of why(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid (CPP-115) isnot expected to cause collateral damage including visual field defects.Some vigabatrin molecules directly inactivate the GABA aminotransferasemolecule upon binding. A small amount of the vigabatrin however, uponbinding to GABA aminotransferase, is converted to 4-oxohex-5-enoic acid,which might diffuse away from the binding site. Due to the reactivity of4-oxohex-5-enoic acid, it could readily bind to other molecules withfunctional groups containing free electron pairs (e.g., NH₂.). Thiscould cause the collateral damage including, but not limited to, visualfield defects and intramyelinic edema.

The mechanism leading to the collateral damage is not known. The damagemight occur from the toxic effects of elevated GABA resulting from theinactivation of GABA-AT, could be a direct toxic effect, could be theconsequence of an enzymatically produced byproduct from the inactivationof GABA-AT, or some combination thereof. The applicants havehypothesized that an alpha-beta unsaturated ketone byproduct resultingfrom the action of GABA-AT on the inactivator molecule.(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid wasspecifically designed to either inactivate the enzyme by an alternatemechanism or to form an intermediate that is much more reactive. Thismore reactive intermediate, if formed, would immediately inactivate theenzyme before it could diffuse into the cytoplasm, thus limiting thepotential for collateral damage to other cellular structures.

Because of the low rate at which vigabatrin actually inactivates theGABA aminotransferase enzyme, vigabatrin is given in higher doses whichresults in 4-oxohex-5-enoic acid being present in higher quantities withgreater opportunity to cause collateral damage.

In contrast, it is believed that the(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid uponoxidation by the GABA aminotransferase immediately inactivates theenzyme because of the highly reactive di-fluoromethylene intermediate,which results from administration of(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid. It ishypothesized that (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoicacid is more thermodynamically favorable for both staying bound to thebinding site and for inducing the conformational changes in GABAaminotransferase upon inactivation of the enzyme. Collateral damage isbelieve to be avoided because the(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid upon amineoxidation immediately inactivates the GABA aminotransferase and remainsbound to the enzyme. Because the reactive intermediate remains bound tothe molecule, it is not free to react with other molecules and thereforedoes not produce the collateral damage. Further, because(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid will be dosedat levels that are more than 100 times lower than vigabatrin, thepotential number of reactive intermediate species is significantlyreduced.

Visual field Experiments

Experiments were conducted to test the hypothesis that(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid reducesvisual field defects.

Materials and Methods

Forty-five male and female Wistar Albino rats (Charles RiverLaboratories), 9 weeks of age at the start of dosing, were acclimated,and placed into one of three treatment groups (vehicle,(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid (CPP-115) orVigabatrin). Animals received a single intra-peritoneal injection ofvehicle or test formulations once daily for either 45 or 90 consecutivedays at 0, 20 or 200 mg/kg for the vehicle,(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid (CPP-115) andVigabatrin treatment groups, respectively. The formulations for eachtreatment group were prepared fresh weekly in 0.9% normal saline.

All animals were acclimated for 7 days to the test facility prior to thestart of dosing and were housed in individual polycarbonate cages. Thecage environment uses a standard 12 hr/12 hr light dark cycle, withstandard industrial fluorescent lighting during the light cycle. Duringthe course of the study, the animals were monitored for mortality,moribundity, clinical signs of illness, feed intake, and body weightchange. Any animals found in distress for more than 24 hours werehumanely euthanized. Due to the sedative effects of both(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid (CPP-115) andVigabatrin, some animals received special food supplements and/or IPfluids in the first 3 weeks of dosing. By the end of the third week ofthe dosing, the sedative effects of both drugs decreased, and no specialfood supplements or IP fluids were required. At the conclusion of thedosing phase (5 of each sex for 45 days and 10 of each sex for 90 days),the animals entered a 5-7 day “washout” period, after whichelectroretinograms (ERGs) were measured for both eyes. The animals werethen humanely euthanized, by CO2 asphyxiation in accordance with AVMAguidelines on euthanasia, for post-mortem pathology examinations.

Electroretinogram Recordings

Following a dark adaptation period of at least 12 hours, each eye wasdilated with tropicamide (1 drop of 1% solution) and phenylepherine (1drop of 10% solution), an anesthetic dose of Ketamine HCl (up to 55mg/kg) and Xylazine HCl (up to 12 mg/kg) was administered IM or IP, andjust prior to the ERG, a topical anesthesia (0.5% Proparacaine HCl) wasapplied to the eye. The types of electroretinogram measurements made andthe electroretinogram testing parameters are found in Table 4.

TABLE 4 Flash Pulse ERG type Flashes/interval Mode Period Intensity Rod5 @ 500 ms Pulse 4 ms 0.02 cd · s/m² Standard 5 @ 40000 ms Pulse 4 ms 7cd · s/m² Combined Light Adaptation 0 @ 900 sec N/A N/A 25 cd/m² SingleFlash - 5 @ 15000 ms Pulse 4 ms 6 cd · s/m² Cone 10 Hz Flicker 20Contin- 4 ms 3 cd · s/m² uous 15 Hz Flicker 20 Contin- 4 ms 3 cd · s/m²uous

The electroretinograms were measured with an Espion² ERG system withColorDome Ganzfeld type illuminator by Diagnosys. All light stimuli werewhite light. The ground electrode was a Grass needle electrode insertedsub-cutaneously at the base of the tail. The reference electrode was aGrass gold disc electrode placed on the tongue. The eye electrode was agold wire. Data was collected on a dual channel system (1 channel pereye) at 1000 samples per second with a 2^(nd) order 0.3 Hz to 300 Hzband pass Bessel filter and also a 60 Hz notch filter to removeinterference from the line power.

Results

The results are shown for the right eye in Table 5 below and for theleft eye in Table 6. The combined data are summarized in FIG. 5. At themaximum tolerated doses,(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid caused muchless damage and had a much larger margin between the therapeutic dosefor addiction and the maximum tolerated dose when compared tovigabatrin. These data clearly show that(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid causes lessvisual field defects than vigabatrin. For the treatment of chronicdisorders, risk of visual field defects may be reduced or eliminated byadministration of (1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoicacid. This data is even more compelling given the fact that thepredicted therapeutic dose of CPP-115 is one twentieth of the dosagegiven in this experiment. The predicted therapeutic dose of CPP-115 is 1mg/kg. Vigabatrin however was dosed at its therapeutic dosage.

TABLE 5 Right Eye Data OD A-wave amplitude A-wave Implicit Time B-waveamplitude Std Std Std Std Std Std Group Rods Combined Cone Rods CombinedCone Rods Combined Cone Control 26.954 121.16 6.2527 25.7 10.2 11.6296.99 428.6 64.026 CPP-115 32.53 99.93125 2.203125 27.875 9.75 10233.8125 313.5375 59.97875 Vigab 32.3765 81.30125 4.744 27.75 18.37512.125 180.35125 276.425 59.69125 Diff from Control CPP-115 5.576−21.22875 −4.049575 2.175 −0.45 −1.6 −63.1775 −115.0625 −4.04725 Vigab5.4225 −39.85875 −1.5087 2.05 8.175 0.525 −116.63875 −152.175 −4.33475B-wave Implicit Time OP Average Std Std Std 10 hz 15 hz Group RodsCombined Cone Combined Flicker Flicker Control 70.6 66.4 42.4 17.411854.978 38.54 CPP-115 72.25 66 42.75 11.77125 47.43 33.11575 Vigab 92.7574.5 52.125 7.053 27.74975 21.201 Diff from Control CPP-115 1.65 −0.40.35 −5.64055 −7.548 −5.42425 Vigab 22.15 8.1 9.725 −10.3588 −27.22825−17.339

TABLE 6 Left Eye Data OS A-wave amplitude A-wave Implicit Time B-waveamplitude Std Std Std Std Std Std Group Rods Combined Cone Rods CombinedCone Rods Combined Cone Rods Control 27.1097 102.762 7.0849 26.1 10.612.1 267.88 361.52 72.3 70.2 CPP-115 37.06625 107.88125 4.380625 28.59.75 14.25 261.4375 335.9 55.5825 71.5 Vigab 24.284625 66.0375 3.73762528.75 18.75 12.375 151.20125 219.4775 40.1925 89 Diff from ControlCPP-115 9.95655 5.11925 −2.70428 2.4 −0.85 2.15 −6.4425 −25.62 −16.71751.3 Vigab −2.825075 −36.7245 −3.34728 2.65 8.15 0.275 −116.67875−142.0425 −32.1075 18.8 B-wave Implicit Time OP Average Std Std Std 10hz 15 hz Group Combined Cone Combined Flicker Flicker Control 67.4 4514.984 56.662 39.538 CPP-115 77.625 43.5 12.172125 42.79875 29.187 Vigab77.5 47.625 6.367625 19.072125 14.8 Diff from Control CPP-115 10.225−1.5 −2.811875 −13.86325 −10.351 Vigab 10.1 2.625 −8.616375 −37.589875−24.738

The disclosure herein is not intended to be limiting and one of skill inthe art will recognize that there are other disorders involving GABAthat can be treated by the compounds and methods of the presentinvention.

We claim:
 1. A method of treating a condition selected from aneurological disorder or a psychological disorder comprisingadministering a gamma-amino butyric acid (GABA) aminotransferaseinhibitor which does not prevent the reuptake of GABA.
 2. The method ofclaim 1 wherein the gamma-amino butyric acid transferase inhibitorinactivates GABA aminotransferase through a di-fluoromethyleneintermediate instead of 4-oxohex-5-enoic acid.
 3. The method of claim 2wherein the GABA aminotransferase inhibitor is(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid or its salts.4. The method of claim 1 where in the treatment has reduced collateraldamage when compared to the administration of an effective amount ofvigabatrin.
 5. The method of claim 4 wherein the collateral damage thatis reduced is selected from visual field defects and intramyelinicedema.
 6. The method of claim 3 wherein the(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid or its saltsis administered at a dose from 0.05 to 2.5 mg/kg/day or from about 3.5to about 175 mg/day for an average adult.
 7. The method of claim 1wherein the patient suffers from a psychological disorder selected fromone or more of the following: general anxiety disorder, pathological orcompulsive gambling disorder, compulsive eating, body dysmorphicdisorder, hypochondriasis, pathologic grooming conditions, kleptomania,pyromania, attention deficit hyperactivity disorder and impulse controldisorders.
 8. The method of claim 1 wherein the neurological disorder isselected from epilepsy, fibromyalgia, neuropathic pain, migraine relatedto epilepsy, restless leg syndrome and post traumatic stress disorderaddiction, obesity, obsessive-compulsive disorders and Alzheimer'sdisease.
 9. A method of treating a psychological disorder or aneurological disorder comprising: administering a gamma-amino butyricacid (GABA) aminotransferase inhibitor which inactivates GABAaminotransferase through a di-fluoromethylene intermediate instead of4-oxohex-5-enoic acid.
 10. The method of claim 9 wherein the gamma-aminobutyric acid transferase gamma-amino butyric acid (GABA)aminotransferase inhibitor does not prevent the reuptake of GABA. 11.The method of claim 10 wherein the GABA aminotransferase inhibitor is(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid or its salts.12. The method of claim 9 where in the treatment has reduced collateraldamage when compared to the administration of an effective amount ofvigabatrin.
 13. The method of claim 12 wherein the collateral damagethat is reduced is selected from visual field defects and intramyelinicedema.
 14. The method of claim 10 wherein the(1S,3S)-3-amino-4-difluoromethylenyl-1-cyclopentanoic acid or its saltsis administered at a dose from 0.05 to 2.5 mg/kg/day or from about 3.5to about 175 mg/day for an average adult.
 15. The method of claim 9wherein the psychological disorder is selected from one or more of thefollowing: general anxiety disorder, pathological or compulsive gamblingdisorder, compulsive eating, body dysmorphic disorder, hypochondriasis,pathologic grooming conditions, kleptomania, pyromania, attentiondeficit hyperactivity disorder and impulse control disorders.
 16. Themethod of claim 12 wherein the neurological disorder is selected fromepilepsy, fibromyalgia, neuropathic pain, migraine related to epilepsy,restless leg syndrome and post traumatic stress disorder addiction,obesity, obsessive-compulsive disorders and Alzheimer's disease.